Long-term objective of this proposal is to understand the structural details of the interaction between the chemokine MIP-Ia and its binding partners, particularly CCR5, its natural receptor. CCR5 is a 7- transmembrane helix receptor that is also used as a co-receptor by HIV in order to gain entry to the cell. I MIP-1beta has been shown to block HIV entry to the cell by binding to CCR5, thereby suppressing HIV infection. In Specific Aim 1 a binding surface used by MIP-1beta to interact with CCR5 is hypothesized, and experiments to define the details of MIP-Ibeta-CCR5 interaction are proposed. In order to elucidate specific properties required for inhibition of HIV, MIP-1beta variants with differing quatemary structure and receptor binding capability will be tested as HIV inhibitors. In Specific Aim 2 NMR structural studies of the interaction between MIP- 113and heparin will be carried out to define the glycosaminoglycan binding surface of MIP- 1131 and to define the quaternary arrangement of MIP-1beta as it binds these sugars. In Specific Aim 3, two strategies will be pursued to investigate possible soluble proxies for the CCR5 receptor. First, X-ray crystallography and NMR spectroscopy will be used to determine the structure of the complex between MIP-1beta. and the virally encoded chemokine binding protein, p35. This will allow both an understanding of this particular complex and may also provide insight on modes of chemokine binding used by the natural receptor CCR5. Second, a novel strategy to investigate the structure of CCR5 is proposed, in which portions of CCR5 are grafted onto a stable scaffold. These three Specific Aims will delineate the molecular details of MIP-1beta binding to its biological partners, and will allow the design of specific, tight-binding anti-HIV therapeutics.